Patients with diabetes commonly display aberrant urogenital (voiding dysfunction) and gastrointestinal (Gl) function (constipation and diarrhea), which may be caused, in part, by autonomic neuropathic changes and altered smooth muscle contractility. A major neurotransmitter influencing contractility is acetylcholine (ACh), which acts via M2 and M3 muscarinic receptors (mAChRs) in smooth muscle. The M3 mAChR mediates direct contraction, whereas the M2 mAChR has been shown to enhance MS-mediated contraction and to inhibit relaxation mediated by the Gs-linked li-adrenoceptor in ileum, colon and bladder. M2 receptor-mediated inhibition of contraction is thought to involve the inhibition of agoniststimulated cAMP accumulation. Although adrenergic innervation is minor in the human bladder, enhanced sympathetic profile in diabetes warrants the study of this mechanism in the bladder. Alterations in M2 and M3 mAChRs and G-adrenergic receptors are known, but there is limited information on their interaction in diabetes. The broad, long-term objective of this investigation is to study how diabetes affects the functional summation of and interplay between muscarinic and other (e.g. adrenergic) receptors in the Gl and urogenital systems, including the indirect contractile role of the M2 receptor, which seems central to these receptor interactions. To fulfill the general objective, muscarinic contractile mechanisms in Gl and urinary bladder smooth muscle from rats and wild type and muscarinic receptor knock out mice rendered diabetic by streptozotocin (STZ) treatment will be investigated. First, changes in postjunctional muscarinic contractility will be measured in isolated smooth muscle from STZ-treated rodents. Second, M3 mAChRs will be eliminated pharmacologically or genetically and the ability of the M2 receptor to inhibit the effects of relaxant agents will be measured. Third, muscarinic receptor-specific signaling assays will be used to assess the effects of diabetes on phosphoinositide hydrolysis (M3) and inhibition of cAMP levels (M2). Fourth, fieldstimulation will be used to determine whether and how the indirect contractile role of the M2 mAChR occurs with neurogenic ACh in diabetes. Characterization of the M2 role may lead to novel therapeutic approaches to modulate and modify receptor interactions to minimize clinical symptoms in diabetes. PUBLIC HEALTH RELEVANCE: A total of 20.8 million people or 7% of the population in the United States have diabetes. Diabetes-related nerve damage may cause abnormal contraction of smooth muscle tissue, such as in the bladder or colon, which may contribute to problems with urination and/or digestion. Studying how diabetes affects the urinary and digestive systems is of considerable importance as it may lead to new treatments.